Dexamethasone protection from TNF-alpha-induced cell death in MCF-7 cells requires NF-kappaB and is independent from AKT

BACKGROUND: The biochemical bases for hormone dependence in breast cancer have been recognized as an important element in tumor resistance, proliferation and metastasis. On this respect, dexamethasone (Dex) dependent protection against TNF-alpha-mediated cell death in the MCF-7 cell line has been demonstrated to be a useful model for the study of this type of cancer. Recently, cytoplasmic signaling induced by steroid receptors has been described, such as the activation of the PI3K/Akt and NF-kappaB pathways. We evaluated their possible participation in the Dex-dependent protection against TNF-alpha-mediated cell death. RESULTS: Cellular cultures of the MCF-7 cell line were exposed to either, TNF-alpha or TNF-alpha and Dex, and cell viability was evaluated. Next, negative dominants of PI3K and IkappaB-alpha, designed to block the PI3K/Akt and NF-kappaB pathways, respectively, were transfected and selection and evaluation of several clones overexpressing the mutants were examined. Also, correlation with inhibitor of apoptosis proteins (IAPs) expression was examined. Independent inhibition of these two pathways allowed us to test their participation in Dex-dependent protection against TNF-alpha-cytotoxicity in MCF-7 cells. Expression of the PI3K dominant negative mutant did not alter the protection conferred by Dex against TNF-alpha mediated cell death. Contrariwise, clones expressing the IkappaB-alpha dominant negative mutant lost the Dex-conferred protection against TNF-alpha. In these clones degradation of c-IAP was accelerated, while that of XIAP was remained unaffected. CONCLUSION: NF-kappaB, but not PI3K/Akt activation, is required for the Dex protective effect against TNF-alpha-mediated cell death, and correlates with lack of degradation of the anti-apoptotic protein c-IAP1

Glycosylated VCAM-1 isoforms revealed in 2D western blots of HUVECs treated with tumoral soluble factors of breast cancer cells

<p>Abstract</p> <p>Background</p> <p>Several common aspects of endothelial phenotype, such as the expression of cell adhesion molecules, are shared between metastasis and inflammation. Here, we analyzed VCAM-1 variants as biological markers of these two types of endothelial cell activation. With the combination of 2-DE and western blot techniques and the aid of tunicamycin, we analyzed N-glycosylation variants of VCAM-1 in primary human endothelial cells stimulated with either TNF or tumoral soluble factors (TSF's) derived from the human breast cancer cell line ZR75.30.</p> <p>Results</p> <p>Treatments induced a pro-adhesive endothelial phenotype. 2D western blots analysis of cells subjected to both treatments revealed the expression of the two known VCAM-1 isoforms and of previously unknown isoforms. In particular TSFZR75.30 induced an isoform with a relative molecular mass (Mr) and isoelectric point (p<it>I</it>) of 75-77 kDa and 5.0, respectively.</p> <p>Conclusion</p> <p>The unknown isoforms of VCAM-1 that were found to be overexpressed after treatment with TSF's compared with TNF, could serve as biomarkers to discriminate between inflammation and metastasis. 2D western blots revealed three new VCAM-1 isoforms expressed in primary human endothelial cells in response to TSF stimulation. Each of these isoforms varies in Mr and pI and could be the result of differential glycosylation states.</p

Efecto anti-tumoral de dos triterpenos aislados de una planta medicinal

Los ácidos masticadienónico (1) y 3α-OH masticadienoico (2) son los principales metabolitos secundarios aislados de la corteza de Amphipterygium adstringens. En trabajos previos se han investigado las propiedades biológicas de estos ácidos. Inicialmente, se demostró la actividad antiinflamatoria en dos modelos de inflamación aguda, carragenina y TPA (12-O-tetradecanoilphorbol-13- acetato) (Oviedo, 2004). Por otro lado, se demostró que estos compuestos inhiben la proliferación de las líneas celulares de cáncer humano (Oviedo, 2005). Respecto al mecanismo anti-tumoral recientemente se informó el efecto de 2 sobre las funciones bioenergéticas y la permeabilidad de la membrana mitocondrial (Dalla, 2012). Sin embargo, no se ha investigado su actividad anti-tumoral in vivo. En este trabajo se investigó el efecto de 1 y 2 sobre el crecimiento tumoral en un modelo murino. Los resultados de esta investigación demuestran que los ácidos 1 y 2 tienen un efecto anti-tumoral en xenotransplantes de carcinoma prostático.The masticadienonic acid (1) and 3α-OH masticadienoic acid (2) are the main secondary metabolites of the barck from Amphipterygium adstringens (Navarrete, 2006). In previous works, were investigated the biological properties of these acids. First, was demonstrated their anti-inflamatory activity by TPA (12-Otetradecanoylphorbol- 13-acetate) and carrageenan inflammatory acute models. Also, has been demonstrated that 1 and 2 compounds inhibit the proliferation of the human cancer cell lines, HCT-15 (colon), MCF-7 (breast), U-251 (CNS), PC-3 (prostate), K-562 (leukaemia), (Oviedo, 2005). Respect to the anti-tumor mechanism recently was informed the effect of 2 on bioenergetics functions and permeability of mithocondrial membrane (Dalla, 2012). However, there are no studies about anti-tumor activity of 1 and 2 in vivo. In this work we investigated the effect of 1 and 2 on growth tumor in a murine model. The results of this research showed that acids 1 and 2 have anti-tumor effect in xenografts of prostatic carcinoma

Establishment of triple-negative breast cancer cells based on BMI: A novel model in the correlation between obesity and breast cancer

IntroductionObesity has been associated with an increased risk of biologically aggressive variants in breast cancer. Women with obesity often have tumors diagnosed at later stages of the disease, associated with a poorer prognosis and a different response to treatment. Human cell lines have been derived from specific subtypes of breast cancer and have served to define the cell physiology of corresponding breast cancer subtypes. However, there are no current cell lines for breast cancer specifically derived from patients with different BMIs. The availability of those breast cancer cell lines should allow to describe and unravel functional alterations linked to these comorbidities. MethodsCell cultures were established from tumor explants. Once generated, the triple negative subtype in a patient with obesity and a patient with a normal BMI were chosen for comparison. For cellular characterization, the following assays were conducted: proliferation assays, chemo – sensitivity assays for doxorubicin and paclitaxel, wound healing motility assays, matrix invasion assays, breast cancer cell growth to estradiol by chronic exposure to leptin, induction of endothelial permeability and tumorigenic potential in athymic mice with normo - versus hypercaloric diets with an evaluation of the epithelium – mesenchymal transformation proteins.ResultsTwo different cell lines, were established from patients with breast cancer: DSG-BC1, with a BMI of 21.9 kg/m2 and DSG-BC2, with a BMI of 31.5 kg/m2. In vitro, these two cell lines show differential growth rates, motility, chemosensitivity, vascular permeability, response to leptin with an activation of the JAK2/STAT3/AKT signaling pathway. In vivo, they displayed distinct tumorigenic potential. In particular, DSG-BC2, presented higher tumorigenicity when implanted in mice fed with a hypercaloric diet.DiscussionTo our knowledge, these primary cultures are the first in vitro representation of both breast cancer and obesity. DSG – BC2 presented a more aggressive in vivo and in vitro phenotype. These results support the hypothesis that breast cancer generated in an obese metabolic state may represent a contrasting variant within the same disease. This new model will allow both further comprehension, functional studies and the analysis of altered molecular mechanisms under the comorbidity of obesity and breast cancer

CD8 Cells of Patients with Diffuse Cutaneous Leishmaniasis Display Functional Exhaustion: The Latter Is Reversed, In Vitro, by TLR2 Agonists

Leishmania mexicana (Lm) causes localized (LCL) and diffuse (DCL) cutaneous leishmaniasis. DCL patients have a poor cellular immune response leading to chronicity. It has been proposed that CD8 T lymphocytes (CD8) play a crucial role in infection clearance, although the role of CD8 cytotoxicity in disease control has not been elucidated. Lesions of DCL patients have been shown to harbor low numbers of CD8, as compared to patients with LCL, and leishmanicidal treatment restores CD8 numbers. The marked response of CD8 towards Leishmania parasites led us to analyze possible functional differences between CD8 from patients with LCL and DCL. We compared IFNγ production, antigen-specific proliferation, and cytotoxicity of CD8 purified from PBMC against autologous macrophages (MO) infected with Leishmania mexicana (MOi). Additionally, we analyzed tissue biopsies from both groups of patients for evidence of cytotoxicity associated with apoptotic cells in the lesions. We found that CD8 cell of DCL patients exhibited low cytotoxicity, low antigen-specific proliferation and low IFNγ production when stimulated with MOi, as compared to LCL patients. Additionally, DCL patients had significantly less TUNEL+ cells in their lesions. These characteristics are similar to cellular “exhaustion” described in chronic infections. We intended to restore the functional capacity of CD8 cells of DCL patients by preincubating them with TLR2 agonists: Lm lipophosphoglycan (LPG) or Pam3Cys. Cytotoxicity against MOi, antigen-specific proliferation and IFNγ production were restored with both stimuli, whereas PD-1 (a molecule associated with cellular exhaustion) expression, was reduced. Our work suggests that CD8 response is associated with control of Lm infection in LCL patients and that chronic infection in DCL patients leads to a state of CD8 functional exhaustion, which could facilitate disease spread. This is the first report that shows the presence of functionally exhausted CD8 T lymphocytes in DCL patients and, additionally, that pre-stimulation with TLR2 ligands can restore the effector mechanisms of CD8 T lymphocytes from DCL patients against Leishmania mexicana-infected macrophages

Los costos de la expansión urbana: aproximación a partir de un modelo de precios hedónicos en la Zona Metropolitana del Valle de México

La Zona Metropolitana del Valle de México (ZMVM) ha experimentado una expansión no proporcional al crecimiento poblacional. Entre otras modalidades de poblamiento, la construcción de grandes conjuntos habitacionales periféricos propicia esa expansión, imponiendo costos individuales y sociales. Se revisan aquí las características de la expansión urbana en la ZMVM y las condiciones de producción de vivienda social periférica, la pérdida de ingreso de los individuos derivada de la distancia a los centros de trabajo y, a través de un modelo de precios hedónicos, se estima el rol de esa distancia en el diferencial de precios entre la vivienda periférica y la intraurbana